The palate separates the oral and nasal cavity and allows breathing to occur alongside eating and drinking. Two broad categories of cell types are critical during palatal development; an epithelial cell layer that covers the palatal shelves and mesenchymal cells that become connective tissue, ultimately providing support and integrity. In humans and mice, dysfunction of either of these cell layers leads to clefting, a common developmental defect with serious postnatal consequences. A cleft can lead to feeding difficulty and the inability to gain or maintain weight at critical neonatal periods. Our group identified Interferon Regulatory Factor 6 (IRF6) as a major cause of clefting in humans. We are interested in identifying the mechanisms of clefting due to mutations in IRF6 and in designing interventions to prevent this birth defect. To realize this goal, our lab developed a mouse model. Thus far, our studies uncovered several important facts about Irf6 and its role in palatal development. First, we know that Irf6 is an essential regulator of proliferation and differentiation in keratinocytes, type of epithelial cell. Second, we demonstrated that Irf6 is expressed most robustly during palatal development in the periderm and Medial Edge Epithelium (MEE), epithelial cells that are critical for palatal formation. Third, we know that changes in Irf6 dose lead to clefting and oral adhesions in the mouse. Finally, we engineered laboratory protocols and mouse models to manipulate the concentration of Irf6 in palatal epithelial cells. Based on these facts, I hypothesize that changes in Irf6 dose in the periderm and MEE will affect palatal development. In Aim I, my goal is to reduce Irf6 dose in the MEE cells of otherwise normal mice and analyze the changes in palatal development that ensue. Based on previous data and preliminary findings, I predict that palatal development will not conclude due to the importance of the MEE and Irf6 in the process. In Aim II, my goal is to deliver Irf6 using an Adenoviral vehicle to periderm cells of Irf6-deficient embryos. In these experiments, I predict that Irf6 delivery to the periderm will rescue cellular function, leading to a significant decrease in oral cavity adhesions. This work has technical, biological and clinical significance. Technically, I have proposed a gene delivery protocol to the oral cavity, something that can be useful to other oral pathologies and genes. We have also created an Irf6 conditional allele, which can be used to assay the function of Irf6 in other tissues and at different timepoints. Biologically, we have proposed highly sophisticated gene manipulation protocols for the periderm and the MEE to examine their specific utility in palatal development as a function of Irf6. Clinically, Aim II provides proof-of principal experiments in mitigating the severity or preventing cleft formation during embryologic development. PUBLIC HEALTH RELEVANCE: Current estimates in the United States suggest that 1/700 individuals are born with a Cleft Lip or Plate (CLP). Patients born with CLP have a greater propensity for multi-system organ dysfunction, including neurological, musculoskeletal, cardiovascular as well as a life- long increased risk of cancer and mortality, each consuming $20,0 from the economy for medical treatment. Studies of palatal development, in health and disease, can lead to rational design of therapeutic interventions and preventative strategies tailored to the patient's genetic footprint.